Detlef Obal

The noble gas xenon induces pharmacological preconditioning in the rat heart in vivo via induction of PKC-ɛ and p38 MAPK

1 Xenon is an anesthetic with minimal hemodynamic side effects, making it an ideal agent for cardiocompromised patients. We investigated if xenon induces pharmacological preconditioning (PC) of the rat heart and elucidated the underlying molecular mechanisms. 2 For infarct size measurements, anesthetized rats were subjected to 25 min of coronary artery occlusion followed by 120 min of reperfusion. Rats received either the anesthetic gas xenon, the volatile anesthetic isoflurane or as positive control ischemic preconditioning (IPC) during three 5‐min periods before 25‐min ischemia. Control animals remained untreated for 45 min. To investigate the involvement of protein kinase C (PKC) and p38 mitogen‐activated protein kinase (MAPK), rats were pretreated with the PKC inhibitor calphostin C (0.1 mg kg−1) or the p38 MAPK inhibitor SB203580 (1 mg kg−1). Additional hearts were excised for Western blot and immunohistochemistry. 3 Infarct size was reduced from 50.9±16.7% in controls to 28.1±10.3% in xenon, 28.6±9.9% in isoflurane and to 28.5±5.4% in IPC hearts. Both, calphostin C and SB203580, abolished the observed cardioprotection after xenon and isoflurane administration but not after IPC. Immunofluorescence staining and Western blot assay revealed an increased phosphorylation and translocation of PKC‐ɛ in xenon treated hearts. This effect could be blocked by calphostin C but not by SB203580. Moreover, the phosphorylation of p38 MAPK was induced by xenon and this effect was blocked by calphostin C. 4 In summary, we demonstrate that xenon induces cardioprotection by PC and that activation of PKC‐ɛ and its downstream target p38 MAPK are central molecular mechanisms involved. Thus, the results of the present study may contribute to elucidate the beneficial cardioprotective effects of this anesthetic gas.

The influence of mitochondrial KATP-channels in the cardioprotection of preconditioning and postconditioning by sevoflurane in the rat in vivo

Volatile anesthetics induce myocardial preconditioning and can also protect the heart when given at the onset of reperfusion—a practice recently termed “postconditioning.” We investigated the role of mitochondrial KATP (mKATP)-channels in sevoflurane-induced cardioprotection for both preconditioning and postconditioning alone and whether there is a synergistic effect of both. Rats were subjected to 25 min of coronary artery occlusion followed by 120 min of reperfusion. Infarct size was determined by triphenyltetrazolium staining. The following protocols were used: 1) preconditioning (S-Pre, n = 10, achieved by 2 periods of 5 min sevoflurane administration (1 MAC) followed by 10 min of washout); 2) sevoflurane postconditioning (1 MAC of sevoflurane given for 2 min at the beginning of reperfusion; S-Post, n = 10); 3) administration before and after ischemia (S-Pre + S-Post, n = 10). Protocols 1–3 were repeated in the presence of 5-hydroxydecanoate (5HD), a specific mKATP-channel-blocker (S-Pre + S-Post + 5HD, S-Pre + 5HD: n = 10; S-Post + 5HD: n = 9). Nine rats served as untreated controls (CON) or received 5HD alone (5HD, n = 10). Both S-Pre (23% ± 13% of the area at risk, mean ± sd) and S-Post (18% ± 5%) reduced infarct size compared with CON (49% ± 11%, both P < 0.05). S-Pre + S-Post resulted in a larger reduction of infarct size (12% ± 5%, P = 0.054 versus S-Pre) compared with administration before or after ischemia alone. 5HD diminished the protection in all three sevoflurane treated groups (S-Pre + 5HD, 35% ± 12%; S-Post + 5HD, 44% ± 12%; S-Pre + S-Post + 5HD, 46% ± 14%;) but given alone had no effect on infarct size (41% ± 13%). Sevoflurane preconditioning and postconditioning protects against myocardial ischemia-reperfusion injury. The combination of preconditioning and postconditioning provides additive cardioprotection and is mediated, at least in part, by mKATP-channels.

Cardiac Myocyte–Specific Expression of Inducible Nitric Oxide Synthase Protects Against Ischemia/Reperfusion Injury by Preventing Mitochondrial Permeability Transition

Background— Inducible nitric oxide synthase (iNOS) is an obligatory mediator of the late phase of ischemic preconditioning, but the mechanisms of its cardioprotective actions are unknown. In addition, it remains unclear whether sustained elevation of iNOS in myocytes provides chronic protection against ischemia/reperfusion injury. Methods and Results— Constitutive overexpression of iNOS in transgenic mice (α-myosin heavy chain promoter) did not induce contractile dysfunction and did not affect mitochondrial respiration or biogenesis, but it profoundly decreased infarct size in mice subjected to 30 minutes of coronary occlusion and 24 hours of reperfusion. In comparison with wild-type hearts, isolated iNOS-transgenic hearts subjected to ischemia for 30 minutes followed by 40 minutes of reperfusion displayed better contractile recovery, smaller infarct size, and less mitochondrial entrapment of 2-deoxy-[3H]-glucose. Reperfusion-induced loss of NAD+ and mitochondrial release of cytochrome c were attenuated in iNOS-transgenic hearts, indicating reduced mitochondrial permeability transition. The NO donor NOC-22 prevented permeability transition in isolated mitochondria, and mitochondrial permeability transition–induced NAD+ loss was decreased in wild-type but not iNOS-null mice treated with the NO donor diethylene triamine/NO 24 hours before ischemia and reperfusion ex vivo. iNOS-mediated cardioprotection was not abolished by atractyloside. Reperfusion-induced production of oxygen-derived free radicals (measured by electron paramagnetic resonance spectroscopy) was attenuated in iNOS-transgenic hearts and was increased in wild-type hearts treated with the mitochondrial permeability transition inhibitor cyclosporin A. Conclusions— Cardiomyocyte-restricted expression of iNOS provides sustained cardioprotection. This cardioprotection is associated with a decrease in reperfusion-induced oxygen radicals and inhibition of mitochondrial swelling and permeability transition.

Desflurane Preconditioning Induces Time-dependent Activation of Protein Kinase C Epsilon and Extracellular Signal-regulated Kinase 1 and 2 in the Rat Heart In Vivo

Background:Activation of protein kinase C epsilon (PKC-&egr;) and extracellular signal-regulated kinase 1 and 2 (ERK1/2) are important for cardioprotection by preconditioning. The present study investigated the time dependency of PKC-&egr; and ERK1/2 activation during desflurane-induced preconditioning in the rat heart. Methods:Anesthetized rats were subjected to regional myocardial ischemia and reperfusion, and infarct size was measured by triphenyltetrazoliumchloride staining (percentage of area at risk). In three groups, desflurane-induced preconditioning was induced by two 5-min periods of desflurane inhalation (1 minimal alveolar concentration), interspersed with two 10-min periods of washout. Three groups did not undergo desflurane-induced preconditioning. The rats received 0.9% saline, the PKC blocker calphostin C, or the ERK1/2 inhibitor PD98059 with or without desflurane preconditioning (each group, n = 7). Additional hearts were excised at four different time points with or without PKC or ERK1/2 blockade: without further treatment, after the first or the second period of desflurane-induced preconditioning, or at the end of the last washout phase (each time point, n = 4). Phosphorylated cytosolic PKC-&egr; and ERK1/2, and membrane translocation of PKC-&egr; were determined by Western blot analysis (average light intensity). Results:Desflurane significantly reduced infarct size from 57.2 ± 4.7% in controls to 35.2 ± 16.7% (desflurane-induced preconditioning, mean ± SD, P < 0.05). Both calphostin C and PD98059 abolished this effect (58.8 ± 13.2% and 64.2 ± 15.4% respectively, both P < 0.05 versus desflurane-induced preconditioning). Cytosolic phosphorylated PKC-&egr; reached its maximum after the second desflurane-induced preconditioning and returned to baseline after the last washout period. Both calphostin C and PD98059 inhibited PKC-&egr; activation. ERK1/2 phosphorylation reached its maximum after the first desflurane-induced preconditioning and returned to baseline after the last washout period. Calphostin C had no effect on ERK1/2 phosphorylation. Conclusions:Both, PKC and ERK1/2 mediate desflurane-induced preconditioning. PKC-&egr; and ERK1/2 are both activated in a time dependent manner during desflurane-induced preconditioning, but ERK1/2 activation during desflurane-induced preconditioning is not PKC dependent. Moreover, ERK1/2 blockade abolished PKC-&egr; activation, suggesting ERK-dependent activation of PKC-&egr; during desflurane-induced preconditioning.

Morphine induces late cardioprotection in rat hearts in vivo: the involvement of opioid receptors and nuclear transcription factor kappaB.

&dgr;1-opioid receptor agonists can induce cardioprotection by early and late preconditioning (LPC). Morphine (MO) is commonly used for pain treatment during acute coronary syndromes. We investigated whether MO can induce myocardial protection by LPC and whether a nuclear transcription factor &kgr;B (NF-&kgr;B)-dependent intracellular signaling pathway is involved. Rats were subjected to 25 min of regional ischemia and 2 h of reperfusion 24 h after treatment with saline (NaCl; 0.9% 5 mL), lipopolysaccharide of Escherichia coli (LPS; 1 mg/kg), or MO (3 mg/kg). LPS is a trigger of LPC and served as positive control. Naloxone (NAL) was used to investigate the role of opioid receptors in LPC and was given before NaCl, LPS, or MO application (trigger phase) or before ischemia-reperfusion (mediator phase). Infarct size (percentage area at risk) was 59% ± 9%, 51% ± 6%, or 53% ± 10% in the NaCl, NAL-NaCl, and NaCl-NAL groups, respectively. Pretreatment with MO reduced infarct size to 20% ± 6% after 24 h (MO-24h), and this effect was abolished by NAL in the trigger (NAL-MO, 53% ± 14%) and in the mediator (MO-NAL, 60% ± 8%) phases. Pretreatment with LPS reduced infarct size to 23% ± 8%. NAL administration in the trigger phase had no effect on infarct size (NAL-LPS 30% ± 16%), whereas NAL during the mediator phase of LPC abolished the LPS-induced cardioprotection (LPS-NAL 54% ± 8%). The role of NF-&kgr;B in morphine-induced LPC was investigated by Western blot and electrophoretic mobility shift assay. Morphine and LPS treatment increased phosphorylation of the inhibitory protein &kgr;B, leading to an increased activity of NF-&kgr;B. Thus, MO induces LPC similarly to LPS and it is likely that this cardioprotection is mediated at least in part by activation of NF-&kgr;B. Opioid receptors are involved as mediators in both MO- and LPS-induced LPC but as triggers only in MO-induced LPC.

Effect of acidotic blood reperfusion on reperfusion injury after coronary artery occlusion in the dog heart.

A prolongation of the intracellular acidosis after myocardial ischemia can protect the myocardium against reperfusion injury. In isolated hearts, this was achieved by prolongation of the extracellular acidosis. The aim of this study was to investigate whether regional reperfusion with acidotic blood after coronary artery occlusion can reduce infarct size and improve myocardial function in vivo. Anesthetized open-chest dogs were instrumented for measurement of regional myocardial function, assessed by sonomicrometry as systolic wall thickening (sWT). Infarct size was determined by triphenyltetrazolium staining after 3 h of reperfusion. The left anterior descending coronary artery (LAD) was perfused through a bypass from the left carotid artery. The animals underwent 1 h of LAD occlusion and subsequent bypass-reperfusion with normal blood (control, n = 6) or blood equilibrated to pH = 6.8 by using 0.1 mM HCl during the first 30 min of reperfusion (HCl, n = 5). Regional collateral blood flow (RCBF) at 30-min occlusion was measured by using colored microspheres. There was no difference in recovery of sWT in the LAD-perfused area between the two groups at the end of the experiments [-2.8+/-1.2% (HCl) vs. -4.4+/-2.5% (control); mean +/- SEM; p = NS]. RCBF was comparable in both groups. Infarct size (percentage of area at risk) was reduced in the treatment group (12.8+/-2.8%) compared with the control group (26.2+/-4.8%; p < 0.05). These results indicate that reperfusion injury after coronary artery occlusion can be reduced by a prolonged local extracellular acidosis in vivo.

Cardioprotection against reperfusion injury is maximal with only two minutes of sevoflurane administration in rats

PurposeVolatile anesthetics can protect the heart against reperfusion injury. When sevoflurane is given for the first 15 min of reperfusion, a concentration corresponding to one minimum alveolar concentration (MAC) provides a maximum protective effect. The present study addresses the question of how long sevoflurane has to be administered to achieve the best cardioprotection.MethodsChloralose anesthetized rats were subjected to a 25-min occlusion of a major coronary artery, followed by 90 min of reperfusion. During the initial phase of reperfusion, an end-tidal concentration of 2.4 vol.% of sevoflurane (1 MAC) was given for two (n = 8), five (n = 8) or ten minutes (n =7). Seven rats served as untreated controls. We measured left ventricular (LV) pressure, mean aortic pressure and infarct size (triphenyltetrazolium staining).ResultsAdministration of sevoflurane for two minutes resulted in the greatest reduction of infarct size to 15% (8–22 [mean (95% confidence interval)] of the area at risk compared with controls [51 (47–55) %, P < 0.00 1], Five or ten minutes of sevoflurane administration reduced infarct size to 26 ( 18–34) and 26 ( 18–35) % [P < 0.05], respectively. The cardiodepressant effect of sevoflurane varied with the duration of its administration: LV dP/dt was reduced from 6332 mmHg · sec−1 (5771–6894) during baseline to 4211 mmHg · sec−1 (3031–5391), 3811 mmHg · sec−1 (2081–5540) and 3612 mmHg · sec−1 (2864–4359) after two, five and ten minutes of reperfusion, respectively.ConclusionAdministration of 1 MAC sevoflurane for the first two minutes of reperfusion effectively protects the heart against reperfusion injury in ratsin vivo. A longer administration time had lesser cardioprotective effects in this experimental model.RésuméObjectifLes anesthésiques voiatiis peuvent protéger le cœur contre tes lésions de reperfusion. Lorsque le sévoflurane est administré pendant les 15 premières minutes de la reperfusion, une concentration correspondant à une concentration alvéolaire minimale (CAM) fournit un effet protecteur maximal. Nous voulions déterminer le temps d’administration de sévoflurane nécessaire pour atteindre la meilleure cardioprotection.MéthodeDes rats anesthésiés à la chloralose ont été soumis à 25 min d’occlusion d’une artère coronaire principale, suivie de 90 min de reperfusion. Pendant la phase initiale de reperfusion, une concentration télé-expiratoire de 2,4 vol.% de sévoflurane (1 CAM) a été donnée pendant deux (n = 8), cinq (n = 8) ou dix minutes (n = 7). Sept rats non traités ont servi de témoins. Nous avons mesuré la pression ventriculaire gauche (VG), la pression aortique moyenne et la taille de l’infarctus (coloration au triphényltétrazolium).RésultatsL’administration de sévoflurane pendant deux minutes a donné la plus importante réduction de la taille de l’infarctus à 15 % de l’aire à risque (8–22 [moyenne (intervalle de confiance de 95 %)], comparativement aux témoins [51 (47–55) %, P < 0,001]. Ladministration pendant 5 à 10 min a respectivement réduit l’infarctus à 26 (18–34) et à 26 (18–35) % [P < 0,05]. L’effet cardiodépresseur du sévoflurane a varié avec la durée de l’administration : la dP/dt du VG a été, respectivement, réduite de 6332 mmHg· secr−1 (5771–6894), comme mesure de base, à 4211 mmHg · sec−1 (3031–5391), 3811 mmHg · sec−1 (2081–5540) et 3612 mmHg · sec−1 (2864–4359) après deux, cinqetdix minutes de reperfusion.ConclusionLadministration de 1 CAM de sévoflurane pendant les deux premières minutes d’une reperfusion protège efficacement le cœur contre les lésions de reperfusion in vivo chez des rats. Une administration prolongée produit moins d’effets cardioprotecteurs chez ce modèle expérimental.

Left stellate ganglion block has only small effects on left ventricular function in awake dogs before and after induction of heart failure.

Left stellate ganglion block (LSGB) results in acute sympathetic denervation of the left ventricular (LV) posterobasal wall. We investigated the effects of LSGB in chronically instrumented awake dogs before and after the induction of pacing-induced congestive heart failure. Twelve dogs were instrumented for measurement of global hemodynamics [LV pressure (LVP)], its first derivative (dP/dt), cardiac output (CO), and regional myocardial function (systolic posterobasal segment length shortening, mean velocity [SLmv]). Before the induction of heart failure (n = 12), LSGB did not affect CO [3.2 ± 1.4 (control, mean ± sd) vs 3.3 ± 1.6 L/min (LSGB, P = 0.45)] and SLmv (11.1 ± 4.0 vs 10.8 ± 4.0 mm/s, P = 0.16), but slightly reduced LVP (130 ± 12 vs 125 ± 14 mm Hg, P = 0.04), dP/dtmax (3614 ± 755 vs 3259 ± 644 mm Hg/s, P = 0.003) and dP/dtmin (−3153 ± 663 vs −2970 ± 725 mm Hg/s, P = 0.03). During heart failure (n = 8), global hemodynamics [CO (2.8 ± 1.2 vs 2.7 ± 1.2 L/min, P = 0.04), LVP (119 ± 6 vs 112 ± 9 mm Hg, P = 0.01), dP/dtmax (1945 ± 520 vs 1824 ± 554 mm Hg/s, P = 0.03) and dP/dtmin (−2402 ± 678 vs −2243 ± 683 mm Hg/s, P = 0.04)], as well as regional myocardial function, were significantly different after LSGB [SLmv] (8.0 ± 3.8 vs 6.9 ± 3.4 mm/s, P = 0.02)]. In conclusion, even during heart failure, the hemodynamic changes after LSGB are small, confirming its broad margin of safety. Implications Left stellate ganglion blockade with local anesthetic produces only very small global hemodynamic and regional myocardial function changes in awake dogs, even in the presence of pacing-induced heart failure.

Genetic Deficiency of Glutathione S-Transferase P Increases Myocardial Sensitivity to Ischemia-Reperfusion Injury

RATIONALE Myocardial ischemia-reperfusion (I/R) results in the generation of oxygen-derived free radicals and the accumulation of lipid peroxidation-derived unsaturated aldehydes. However, the contribution of aldehydes to myocardial I/R injury has not been assessed. OBJECTIVE We tested the hypothesis that removal of aldehydes by glutathione S-transferase P (GSTP) diminishes I/R injury. METHODS AND RESULTS In adult male C57BL/6 mouse hearts, Gstp1/2 was the most abundant GST transcript followed by Gsta4 and Gstm4.1, and GSTP activity was a significant fraction of the total GST activity. mGstp1/2 deletion reduced total GST activity, but no compensatory increase in GSTA and GSTM or major antioxidant enzymes was observed. Genetic deficiency of GSTP did not alter cardiac function, but in comparison with hearts from wild-type mice, the hearts isolated from GSTP-null mice were more sensitive to I/R injury. Disruption of the GSTP gene also increased infarct size after coronary occlusion in situ. Ischemia significantly increased acrolein in hearts, and GSTP deficiency induced significant deficits in the metabolism of the unsaturated aldehyde, acrolein, but not in the metabolism of 4-hydroxy-trans-2-nonenal or trans-2-hexanal; on ischemia, the GSTP-null hearts accumulated more acrolein-modified proteins than wild-type hearts. GSTP deficiency did not affect I/R-induced free radical generation, c-Jun N-terminal kinase activation, or depletion of reduced glutathione. Acrolein exposure induced a hyperpolarizing shift in INa, and acrolein-induced cell death was delayed by SN-6, a Na(+)/Ca(++) exchange inhibitor. Cardiomyocytes isolated from GSTP-null hearts were more sensitive than wild-type myocytes to acrolein-induced protein crosslinking and cell death. CONCLUSIONS GSTP protects the heart from I/R injury by facilitating the detoxification of cytotoxic aldehydes, such as acrolein.

Ultrasound guidance for deep peripheral nerve blocks: a brief review.

Nerve stimulation and ultrasound have been introduced to the practice of regional anesthesia mostly in the last two decades. Ultrasound did not gain as much popularity as the nerve stimulation until a decade ago because of the simplicity, accuracy and portability of the nerve stimulator. Ultrasound is now available in most academic centers practicing regional anesthesia and is a popular tool amongst trainees for performance of nerve blocks. This review article specifically discusses the role of ultrasonography for deeply situated nerves or plexuses such as the infraclavicular block for the upper extremity and lumbar plexus and sciatic nerve blocks for the lower extremity. Transitioning from nerve stimulation to ultrasound-guided blocks alone or in combination is beneficial in certain scenarios. However, not every patient undergoing regional anesthesia technique benefits from the use of ultrasound, especially when circumstances resulting in difficult visualization such as deep nerve blocks and/or block performed by inexperienced ultrasonographers. The use of ultrasound does not replace experience and knowledge of relevant anatomy, especially for visualization of deep structures. In certain scenarios, ultrasound may not offer additional value and substantial amount of time may be spent trying to find relevant structures or even provide a false sense of security, especially to an inexperienced operator. We look at available literature on the role of ultrasound for the performance of deep peripheral nerve blocks and its benefits.